The mammalian brain is vigilant in control of breathing, regulating blood O2 and CO2 over a wide range of metabolic demand from birth till death. The brain makes efficient use of the respiratory musculature because the metabolic cost of continually breathing is considerable. In our attempt to understand the action of the brain in breathing, we ask two questions: How is respiratory rhythm generated? and, How is this rhythm transformed into a precisely modulated pattern of respiratory muscle activity? We address the first question in this proposal. In order to identify the neural mechanism underlying rhythmogenesis, one must first establish which neurons are involved. We hypothesize that neurons with pacemaker properties in the preBotzinger Complex (preBOtC) in the rostral ventrolateral medulla underlite rhythmogensis. The crux of testing the hypothesis that preBOtC pacemaker neurons are the kernel for generating respiratory rhythm (pacemaker hypothesis) is testing the causal role, if any, of pacemaker cells in rhythm generation. We propose to test the pacemaker hypothesis by: characterizing the ionic basis for bursting in pacemaker cells of the preBOtC. Determining if pacemaker neurons have the predicted responses to extrinsic stimuli that reset the respiratory phase timing. Determining if pacemaker neurons have the obligatory synaptic interactions if they are the source of respiratory rhythm. Determination of the mechanisms underlying breathing movements is basic to understanding human physiology and the pathophysiology of many diseases. Development of prophylaxis and treatment of such diseases as Sudden Infant Death Syndrome, apnea of prematurity, central alveolar hypoventilation, congenital central hypoventilation syndrome, sleep apnea and other forms of respiratory failure critically depend on such knowledge. Furthermore, the proposed work provides a unique exploration of an important integrative action of the brain. Exploiting a measurable behavior under controlled in vitro conditions permits novel experiments that can reveal important features in the link between synapses/neurons and behavior.